The interleukin-2 receptor and related cytokine/cytokine receptor systems are being studied to understand critical components of the T cell immune response in normal and neoplastic cells. Following T-cell activation, IL-2 and IL-2 receptors are induced; the magnitude and duration of the T-cell immune response is controlled by the amount of IL-2 produced, the levels of receptors expressed, and the time course of these events. Expression of IL-2Ra is interestingly high in cells infected with HTLV-I, the cause of adult T cell leukemia (ATL) and tropical spastic paraparesis/HTLV-I-associated myelopathy (TSP/HAM). Three chains of the IL-2 receptor exist, IL-2Ra, IL-2Rb, and gc, with IL-2Ra and IL-2Rb being significantly regulated at the level of transcription. gc is a shared chain also used by the receptors for IL-4, IL-7, IL-9, IL-15, and IL-21, and is the protein that is mutated in XSCID. We have focused primarily on the types of signals induced by some of these cytokines, particularly the activation of STAT proteins (signal transducers and activators of transcription), and the mechanism by which they regulate cytokine/STAT target genes. Given our prior observations that STAT5A or STAT5B transgenic mice develop tumors, which was consistent with STAT5 being implicated in malignant transformation and elevated in a range of human tumors, this is an important area for both normal and pathological states. Moreover, humans and mice with defective STAT protein expression have a range of immunological defects. In the past year, in a collaboration, we reported that enhanced T cell lymphoma in NOD-Stat5b transgenic mice is caused by hyperactivation of STAT5B in CD8+ T cells. T helper cell differentiation is critical for normal immune responses, with Th1 differentiation being important for host defense to viruses and other intracelllular pathogens, Th2 differentiation being vital in allergic disorders and related to helminths, and Th17 differentiation being vital in a range of inflammatory disorders, including psoriasis and inflammatory bowel disease. We previously showed that IL-2 is important for Th2 differentiation and reported that IL-2 regulates expression of the IL-4 receptor in a STAT5-dependent manner and critically controls priming of cells for Th2 differentiation. Moreover, using genome-wide Ilumina-based ChIP-Seq (chromatin immunoprecipitation coupled to DNA sequencing) analysis, we previously discovered broad regulation of Th2 differentiation via STAT5A and STAT5B, substantially extending earlier studies focused on STAT5A. Moreover, we had discovered that IL-2-mediated IL-4Ra induction was critical in priming cells for Th2 differentiation. In the prior year, we substantially extended these findings by showing that IL-2 via STAT5 induces expression of IL-12Rb1 and IL-12Rb2 and that the induction of IL-12Rb2 is critical for Th1 differentiation and we defined the mechanism of regulation of IL-12Rb2. Additionally, we showed that IL-2 via STAT5 also regulates the T box protein, T-bet. Interestingly, in contrast to the induction of IL-12R proteins, IL-2 inhibits expression of IL-6Ra and gp130, helping to explain the inhibition of Th17 differentiation. Consistent with the ability of Tbx21 to inhibit Th17 differentiation, expression of Tbx21 in Th17 cells resulted in increased IFNg but decreased expression of IL-17A. These results indicated a very broad effect of IL-2 via STAT5 on T helper cell differentation. In the current review year, we have continued to study the role of IL-2 in Th differentiation, including related to Th9 differentiation. During the past year, we also continued our collaboration with Dr. K. Christopher Garcia at Stanford, studying the actions of wild type IL-2 versus novel IL-2 variants, a project with potential clinical ramifications. These studies in part use the pmel-1 T cell receptor transgenic model of adoptive immunotherapy for cancer in collaboration with Dr. Nicholas Restifo, NCI. We have also collaborated with Dr. Garcia on a project in which the three dimensional structure of IL-2 complexed to its receptor was compared to that of IL-15 bound to its receptor. These studies, reported in the past year, have provided key mechanistic and structural insights into the functional differences between IL-2 and IL-15, which are highly related and share IL-2Rbeta and gc as receptor components but nevertheless possess distinctive biological functions. Although IL-2 primarily signals via cis-signaling and IL-15 via trans-signaling, these cytokines have essentially identical activation of STAT, PI3K/Akt, and Ras/MAPK signaling pathways. Moreover, gene expression profiles are very similar, although not identical. Thus, these cytokines have almost indistinguishable signaling properties despite different biological responses. This study has substantially elucidated structural and mechanistic aspects of IL-2 and IL-15 signaling. Previously, we demonstrated that IL-21 regulated expression of the Prdm1 gene that encodes BLIMP1 via a response element that depends on STAT3 and IRF4. This led to our reporting in the past year that in contrast to its known ability to cooperate with PU.1 in B cells to act via Ets-IRF composite elements (EICEs), IRF4 cooperates with BATF/JUN family proteins to act via AP1-IRF composite elements (AICEs) in T cells, as well as in B cells. We demonstrated critical cooperative regulation of important genes via these AICEs and demonstrated cooperative binding of IRF4, BATF, and JUN family proteins, with markedly diminished IRF4 binding in Batf-deficient cells and markedly diminished BATF binding in Irf4-deficient cells. We demonstrated critical regulation of key genes, including for example those encoding IL-10 and IL-17 via AICEs. In collaborative studies with Ken Murphy, we also reported that there were important compensatory roles for BATF factors in dendritic cell development mediated by BATF-IRF interactions involving the leucine zipper domain of BATF. We also reported key actions of cross-talk between IL-21/STAT3 and GM-CSF/STAT5. IL-21 has broad actions on T- and B-cells, but its actions in innate immunity are poorly understood. We reported that IL-21 induced apoptosis of conventional dendritic cells (cDCs) via STAT3 and Bim, and this was inhibited by granulocyte-macrophage colony-stimulating factor (GM-CSF). ChIP-Seq analysis revealed genome-wide binding competition between GM-CSF-induced STAT5 and IL-21-induced STAT3. Expression of IL-21 in vivo decreased cDC numbers, and this was prevented by GM-CSF. Moreover, repetitive &#945;-galactosylceramide injection of mice induced IL-21 but decreased GM-CSF production by natural killer T (NKT) cells, correlating with decreased cDC numbers. Furthermore, adoptive-transfer of wild-type CD4+ T cells caused more severe colitis with increased DCs and interferon (IFN)-&#947;-producing CD4+ T cells in Il21r-/-Rag2-/- mice (which lack T cells and have IL-21-unresponsive DCs) than in Rag2-/- mice. Thus, IL-21 and GM-CSF exhibit cross-regulatory actions on gene regulation and apoptosis, regulating cDC numbers and thereby the magnitude of the immune response. Overall, the above findings enhance our understanding of mechanisms by which the gc family cytokines regulate gene expression and biologically important processes. In addition, these findings have implications related to the treatment of cancer, autoimmune, and other diseases.